Optimization of Nucleation and Crystallization for Lyophilization Using Gap Freezing

ABSTRACT

This application discloses devices, articles, and methods useful for producing lyophilized cakes of solutes. The devices and articles provide for a method of freezing liquid solutions of the solute by the top and the bottom of the solution simultaneously and at approximately the same rate. The as frozen solution can then provide a lyophilized cake of the solutes with large and uniform pores.

CROSS-REFERENCE TO RELATED APPLICATION

This is a continuation of U.S. patent application Ser. No. 14/543,627,filed Nov. 17, 2014, which is a continuation of U.S. patent applicationSer. No. 13/432,498, filed Mar. 28, 2012, which is acontinuation-in-part of U.S. patent application Ser. No. 13/246,342,filed, Sep. 27, 2011, and the benefit under 35 U.S.C. §119(e) of U.S.Provisional Patent Application Ser. No. 61/387,295 filed Sep. 28, 2010,is also hereby claimed. The disclosures of the foregoing applicationsare hereby incorporated herein by reference.

FIELD OF DISCLOSURE

This disclosure relates to methods and apparatus used for lyophilizingliquid solutions of solutes. The disclosure provides a method foroptimization of the nucleation and crystallization of the liquidsolution during freezing to produce lyophilized cakes of the soluteswith large, consistent pore sizes. The disclosure also provides a methodfor rapid lyophilization of the frozen liquid solution. The disclosureadditionally provides apparatus for use with the method andlyophilization chambers.

BRIEF DESCRIPTION OF RELATED TECHNOLOGY

The preservation of materials encompasses a variety of methods. Oneimportant method, lyophilization, involves the freeze-drying of solutes.Typically, a solution is loaded into a lyophilization chamber, thesolution is frozen, and the frozen solvent is removed by sublimationunder reduced pressure.

One well known issue associated with the lyophilization of materials(e.g., sugars) is the formation of one or more layers of the solute (thedissolved materials) on the top of the frozen solution. In a worse case,the solute forms an amorphous solid that is nearly impermeable andeventually prevents sublimation of the frozen solvent. These layers ofconcentrated solute can inhibit the sublimation of the frozen solventand may require use of higher drying temperatures and/or longer dryingtimes. The higher drying temperatures may negatively impact theintegrity of the solute and the longer drying time may have a negativeeffect on the economics of the process.

SUMMARY

One embodiment of the invention is an article adapted for use in alyophilization chamber comprising a heat sink with a heat sink surfacein thermal communication with a refrigerant; a tray surface; and athermal insulator disposed between the heat sink surface and the traysurface. The article can include a refrigerant conduit in thermalcommunication with the heat sink surface and a heat sink medium disposedbetween the refrigerant conduit and the heat sink surface.

The thermal insulator can form a fixed distance, for example greaterthan about 0.5 mm, separating the heat sink surface and tray surfaceduring one or more steps in a lyophilization process. The distance canbe maintained by the insulator comprising a spacer disposed between theheat sink surface and the tray surface, the spacer having a thickness ofgreater than, for example, about 0.5 mm. In an embodiment the thermalinsulator can support a tray carrying the tray surface. In a furtherembodiment the thermal insulator can form the tray surface.

An additional embodiment of the invention is the lyophilization devicethat includes the article. In this embodiment, the lyophilization devicecan include a plurality of heat sinks that individually have a heat sinksurface in thermal communication with a refrigerant, at least one ofsaid heat sinks being disposed above another to thereby form upper andlower heat sinks; a tray surface disposed between the upper heat sinkand a lower heat sink surface; and a thermal insulator is disposedbetween the tray surface and the lower heat sink.

The lyophilization device can have the distance from the heat sinksurface to the tray surface fixed by the thermal insulator. The thermalinsulator can comprise the spacer, or a brace affixed to an internalwall (fixed or adjustable) of the lyophilization device or otherembodiments can maintain a distance between the lower heat sink surfaceand the tray surface during one or more steps in the lyophilizationprocess.

Still another embodiment of the invention is a vial comprising asealable sample container having top and a bottom and a thermalinsulator comprises a thermally insulating support affixed to the bottomof the sealable sample container, the thermally insulating supporthaving a thermal conductivity less than about 0.2 W/mK at 25° C. Wherethe sample container and the insulating support are made of differentmaterials.

Yet another embodiment is a method of lyophilizing a liquid solutionusing the article, lyophilization device and/or vial described herein.The method includes loading a container comprising a liquid solutioninto a lyophilization chamber comprising a heat sink; the liquidsolution comprising a solute and a solvent and characterized by a topsurface and a bottom surface; providing a thermal insulator between thecontainer and the heat sink; lowering the temperature of the heat sinkand thereby the ambient temperature in the lyophilization chambercomprising the container to a temperature sufficient to freeze theliquid solution from the top and the bottom surfaces at approximatelythe same temperature and form a frozen solution. The method thenincludes lyophilizing the frozen solution by reducing the ambientpressure. In a further embodiment the method may comprise removing thethermal insulator before or during the lyophilizing step.

The method can include the lyophilization chamber having a plurality ofheat sinks and loading the container comprising the liquid solution intothe lyophilization chamber between two parallel heat sinks.

A further embodiment of the invention includes a method of freezing aliquid solution for subsequent lyophilization, the liquid comprising topand bottom surfaces and disposed in a container, and the containerdisposed in a lyophilization chamber comprising a heat sink, theimprovement comprising forming the a thermal insulator by separating thecontainer from direct contact with the heat sink, to thereby freeze thesolution from the top and bottom surfaces at approximately the sametemperature.

Still another embodiment of the invention is a lyophilized cakecomprising a substantially dry lyophilized material; and a plurality ofpores in the lyophilized material having substantially the same poresize; wherein the lyophilized cake was made by the method disclosedherein. The lyophilized cake can have a pore size that is substantiallylarger than the pore size of a reference lyophilized cake comprising thesame material as the lyophilized cake but made by a reference methodcomprising loading a container comprising a liquid solution into alyophilization chamber comprising a heat sink; the liquid solutioncomprising the material and a solvent; excluding a thermal insulatorbetween the container and the heat sink; lowering the temperature of theheat sink and thereby the ambient temperature in the lyophilizationchamber to freeze the liquid solution; freezing the liquid solution; andlyophilizing the frozen solution to form the reference lyophilized cake.

Yet another aspect of the invention is a method including providing alyophilization chamber including a heat sink surface in thermalcommunication with a refrigerant, loading a container including a liquidsolution into the lyophilization chamber, the liquid solution includinga solute and a solvent and characterized by a top surface and a bottomsurface, and lowering the temperature of the heat sink and thereby theambient temperature in the lyophilization chamber holding the containerto a temperature sufficient to freeze the liquid solution, wherein thecontainer is loaded into the lyophilization chamber at a distance spacedvertically from the heat sink thereby forming an intervening thermalinsulator, the distance selected to provide freezing the liquid solutionfrom the top and the bottom surfaces at approximately the sametemperature and thereby form a frozen solution.

In any one of the methods described herein employing a thermal insulatorbetween container and heat sink during freezing, it is furthercontemplated that the thermal insulator may be removed thereby placingthe container in thermally-conductive contact (i.e., direct or indirect)with the heat sink during or following freezing of the solution, tothereby facilitate more rapid freeze drying in the sublimation process.

BRIEF DESCRIPTION OF THE DRAWING FIGURES

For a more complete understanding of the disclosure, reference should bemade to the following detailed description and accompanying drawingfigures wherein:

FIG. 1 is a drawing of the inside of a lyophilization device showing alyophilization chamber and a plurality of heat sinks in a verticalarrangement;

FIG. 2 is a composite drawing of an article showing an arrangement of aheat sink surface and a tray surface;

FIG. 3 is another composite drawing of an article showing an arrangementof a plurality of heat sinks and the location and separation of the heatsink surface and the tray surface;

FIG. 4 is illustrations of sample containers, here vials, (4 a)positioned on a tray, (4 b) positioned directly on a thermal insulator,or (4 c) combined with a thermally insulating support;

FIG. 5 is a drawing of a sample vial including a liquid solution showingthe placement of thermocouples useful for the measurement of thetemperatures of the top and the bottom of the solution;

FIG. 6 is a plot of the temperatures of the top and the bottom of a 10wt. % aqueous sucrose solution frozen using a 3 mm gap between a heatsink surface and a tray (the tray having a thickness of about 1.2 mm)showing a nucleation event, the differences in temperatures between thetop and the bottom of the solution, and the reduction in temperature ofthe top of the solution after the freezing point plateau;

FIGS. 7A and 7B are plots of the water-ice conversion indices for a 5wt. % aqueous sucrose solution as a function of distance (air gap) froma heat sink surface to a tray (the tray having a thickness of about 1.2mm), wherein FIG. 7A plots gap thicknesses over the range of 0-25 mm andFIG. 7B plots gap thicknesses over the range 0-8 mm;

FIG. 8 is a plot of the internal temperatures of vials during a primarydrying process illustrating the effect of gap-freezing on the producttemperature during freeze-drying;

FIG. 9 is a plot of effective pore radii for samples frozen on a 6 mmgapped tray and samples frozen directly on the heat sink surface;

FIGS. 10 and 11 are temperature probe diagrams for top shelf and bottomshelf vials according to Example 2;

FIGS. 12 and 13 are comparisons of approximate drying time for vials onthe top shelf and bottom shelf, according to Example 2;

FIG. 14 is a comparison of product temperatures of top shelf centervials and bottom shelf center vials during drying, according to Example2;

FIG. 15 is a comparison of product temperatures of top shelf centervials and edge vials TP04 and TP07 during drying, according to Example2; and

FIG. 16 is a comparison of product temperatures of bottom shelf centervials and edge vials during drying, according to Example 2.

While the disclosed methods and articles are susceptible of embodimentsin various forms, there are illustrated in the examples and figures (andwill hereafter be described) specific embodiments of the methods andarticles, with the understanding that the disclosure is intended to beillustrative, and is not intended to limit the invention to the specificembodiments described and illustrated herein.

DETAILED DESCRIPTION

One well known issue associated with the lyophilization of materials(e.g., sugars) is the formation of one or more layers of the solute (thedissolved materials) on the top of the frozen solution. These layersform during the freezing of the solution because, typically, thesolutions are positioned within the lyophilization chamber on a heatsink which rapidly decreases in temperature and causes the solution tofreeze from the bottom up. This bottom up freezing pushes the solute inthe liquid phase closer to the top of the solution and increases thesolute concentration in the still liquid solution. The highconcentration of solute can then form a solid mass that can inhibit theflow of gasses therethrough. In a worse case, the solute forms anamorphous solid that is nearly impermeable and prevents sublimation ofthe frozen solvent. These layers of concentrated solute can inhibit thesublimation of the frozen solvent and may require use of higher dryingtemperatures and/or longer drying times.

Disclosed herein is an apparatus for and method of freezing a material,e.g., for subsequent lyophilization, that can prevent the formation ofthese layers and thereby provide efficient sublimation of the frozensolvent.

The lyophilization or freeze drying of solutes is the sublimation offrozen liquids, leaving a non-subliming material as a resultant product.Herein, the non-subliming material is generally referred to as a solute.A common lyophilization procedure involves loading a lyophilizationchamber with a container that contains a liquid solution of at least onesolute. The liquid solution is then frozen. After freezing, the pressurein the chamber is reduced sufficiently to sublime the frozen solvent,such as water, from the frozen solution.

The lyophilization device or chamber is adapted for the freeze drying ofsamples in containers by including at least one tray for supporting thecontainer and means for reducing the pressure in the chamber (e.g., avacuum pump). Many lyophilization devices and chambers are commerciallyavailable.

With reference to FIGS. 1-3, the lyophilization chamber includes a heatsink 101 that facilitates the lowering of the temperature within thechamber. The heat sink 101 includes a heat sink surface 102 that isexposed to the internal volume of the lyophilization chamber and is inthermal communication with a refrigerant 103. The refrigerant 103 can becarried in the heat sink 101 within a refrigerant conduit 104. Therefrigerant conduit 104 can carry the heat sink surface 102 or can be influid communication with the heat sink surface 102 for example through aheat sink medium 105. The heat sink medium 105 is a thermal conductor,not insulator, and preferably has a thermal conductivity of greater thanabout 0.25, 0.5, and/or 1 W/mK at 25° C.

According to the novel method described herein, the sample containers106 do not sit on or in direct, substantial thermal conductivity withthe heat sink 101 during freezing. In one embodiment, the samplecontainers 106 sit on or are carried by a tray surface 107 that isthermally insulated from the heat sink 101. In another embodiment, thesample containers 106 are thermally insulated by being suspended abovethe heat sink 101.

The tray surface 107 is thermally insulated from the heat sink 101 by athermal insulator 108. The thermal insulator 108 has a thermalconductivity of less than about 0.2, less than 0.1, and/or less than0.05 W/mK at 25° C. The thermal insulator 108 can be a gas, a partial orcomplete vacuum, a paper, a foam (e.g., a foam having flexibility atcryogenic temperatures), a polymeric material, or a combination or othermixture of thereof. The polymeric material can be free of orsubstantially free of open cells or can be a polymeric foam (e.g., acured foam). As used herein, the thermal insulator 108 refers to thematerial, object and/or space that provides thermal insulation from theheat sink 101. Air is still considered a thermal insulator in a methodor apparatus wherein the pressure of the air is decreased due toevacuation of the lyophilization chamber.

The level of thermal insulation provided by the thermal insulator 108can be dependent on the thickness of the thermal insulator 108. Thisthickness can be measured by the distance 109 from the heat sink surface102 to the tray surface 107, for example. This distance 109, limited bythe internal size of the lyophilization chamber, can be in a range ofabout 0.5 to about 50 mm, for example, or smaller if the thermalisolation is very high. This distance 109 can be optimized for specificlyophilization chamber volumes and preferably is greater than about 0.5,0.75, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9,9.5, 10, 15, 20, 25, 30, 35, 40, 45, or 50 mm. While the distance 109can be larger than about 10 mm, the volume within the lyophilizationdevice is typically better used by optimizing the distances below about20 mm. Notably, the distance between the heat sink surface 102 and thetray surface 107 is only limited by the distance between the heat sinksurface 102 and the upper heat sink 101 minus the height of a vial 106.The preferred distance 109 can be dependent on the specific model andcondition of lyophilization chamber, heat sink, refrigerant, and thelike, and is readily optimized by the person of ordinary skill in viewof the present disclosure to avoid uneven freezing from top and bottomsurfaces of the solution in the container.

In an embodiment where the tray surface 107 is thermally insulated fromthe heat sink 101, the tray surface 107 is carried by a tray 110,preferably a rigid tray. Notably, the tray surface 107 can be a thermalinsulator (e.g., foamed polyurethane) or a thermal conductor (e.g.,stainless steel). In such an embodiment the thermal insulator 108 maycomprise a gas, a partial vacuum, or a full vacuum.

The tray 110 is preferably maintained at a fixed distance between heatsink surface 102 and the tray surface 107 during freezing. The tray 110can be spaced from the heat sink surface 102 by the thermal insulator108 formed in an embodiment to include a spacer 111 positioned betweenthe tray 110 and the heat sink surface 102 or can be spaced from theheat sink surface 102 to form the thermal insulator 108 by operationallyengaging the tray 110 to a bracket 112 affixed to an internal surface113 (e.g., wall) of the lyophilization chamber. In a further embodiment,the tray 110 is maintained at a distance from the heat sink surface 102to form the thermal insulator 108 by a plurality of struts (not shown)that operationally engage the tray 110 and heat sink surface 102. In anembodiment where a spacer 111 supports the tray 110, the distance fromthe heat sink surface 102 to the tray surface 107 is the thickness ofthe spacer 111 plus the thickness of the tray 110. In agreement with thedistances disclosed above, the spacer 111 can have a thickness in arange of about 0.5 mm to about 10 mm, about 1 mm to about 9 mm, about 2mm to about 8 mm, and/or about 3 mm to about 7 mm, for example. The tray110 can be carried by one or more spacers 111 placed between the heatsink surface 102 and the tray 110.

In another embodiment, the tray 110 can be carried by the thermalinsulator 108 comprising a rigid thermal insulator. For example the tray110 can be a thermal conductor (e.g., stainless steel) and supported by(e.g., resting on) a thermal insulator (e.g., foamed polyurethane). In afurther embodiment the rigid thermal insulator can be combined withspacers to carry the tray. In agreement with the distances disclosedabove, the rigid thermal insulator (with or without the spacer) can havea thickness in a range of about 0.5 mm to about 10 mm, about 1 mm toabout 9 mm, about 2 mm to about 8 mm, and/or about 3 mm to about 7 mm,for example.

The lyophilization device can include a plurality of heat sinks 101 thatindividually have a heat sink surface 102 in thermal communication witha refrigerant 103. In such a lyophilization device, the heat sinks 101can be disposed vertically in the lyophilization chamber with respect toeach other, forming upper and lower heat sinks 101 (see e.g., FIG. 1).By convention, the lower heat sink surface 102 is disposed between theupper and lower heat sinks and the tray surface 107 is disposed betweenthe upper heat sink 101 and the lower heat sink surface 102. In thisarrangement, the thermal insulator 108 is disposed between the traysurface 107 and the lower heat sink 101.

In another embodiment, each individual sample container 106 can sit onor be carried by a thermal insulator 108 (see e.g., FIG. 4 b). Forexample, when the sample container is a vial having a top and a bottomthe thermal insulator 108 can comprise a thermally insulating support114 affixed to the bottom of the vial 115 (see e.g., FIG. 4 c). Thethermally insulating support 114 can have a thermal conductivity lessthan about 0.2 W/mK, less than about 0.1 W/mK, and/or less than about0.05 W/mK at 25° C., for example. In one embodiment, the vial 106 andthe insulating support 114 are different materials (e.g., the vial cancomprise a glass and the insulating support can comprise a foam or apolymer). The vial can comprise a sealable vial.

Another embodiment of the invention includes a method of freezing aliquid solution for subsequent lyophilization. In one embodiment of themethod, the lyophilization chamber as described above is loaded with aliquid solution held in a container that includes a solute (e.g., anactive pharmaceutical agent) and a solvent. The liquid solution willhave a top surface 116 and a bottom surface, wherein the bottom surface117 is proximal to the heat sink 101 (see FIG. 5). The container isseparated from the heat sink 101 by providing a thermal insulator 108between the container and the heat sink 101, the thermal insulatorhaving the characteristics described herein. Thus, the container isspaced vertically from the heat sink with an intervening thermalinsulator 108, the distance and thermal insulator 108 being selected toprovide freezing of the liquid solution from the top and bottom surfacesat approximately the same rate. Having been loaded into thelyophilization chamber, the liquid solution can be frozen by loweringthe temperature of the heat sink 101 and thereby the ambient temperaturein the lyophilization chamber. The liquid solution advantageously can befrozen from the top and the bottom surfaces at approximately the samerate to form a frozen solution. A further advantage is that theconcurrent water to ice conversion at the top and bottom of the solutionavoids problematic freeze-concentration and skin formation observed whenthe bottom of the solution freezes more rapidly than the top.

A further embodiment of the inventions includes once frozen, the liquidsolution (now the frozen solution) can be lyophilized to yield alyophilized cake. In one type of embodiment, the solution is lyophilizedwithout any significant change in the thermal insulator 108 such as byexample maintaining the physical arrangement of the container and heatsink elements. In another type of embodiment, the container having thefrozen liquid solution is placed in thermally-conductive contact withthe heat sink during or following freezing, for example by removing thethermal insulator 108 and placing the tray 107 or containers directly onthe shelf. Embodiments of the removal can comprise, removing the spacer111, moving the brackets 112 or altering the length of the struts (notshown). It is also envisioned that the thermal insulator container notbe placed in thermally conductive contact with the heat sink but theinsulation characteristics of the thermal insulator 108 be altered suchas by significantly lessening the insulation characteristics by reducingthe spacing between the tray and heat sink to a minimal distance.

As noted in connection with Example 2 below, when freezing and drying anarray of containers, containers placed at the edges of such an array,and those especially at the corners, can experience temperatures whichdeviate from those of center containers, due to radiant heat from sidewalls. Thus, in a method of freezing an array of containers, it iscontemplated that the thermal insulator 108 or portions thereof betweenone or more of the container and heat sink can dimensionally vary fromthe thermal insulator or portions thereof between one or more remainingcontainers. In an embodiment, the thickness of the thermal insulator 108can be reduced for edge and/or corner containers, relative to thethickness of the thermal insulator 108 between center containers andheat sink, in order to counter-balance the radiant heating experiencedby such edge and corner containers from side walls and thus achieve moreconsistent temperature profiles across the array.

In this embodiment, the thermal insulator provides for the facilefreezing of the liquid solution from the top and the bottom within thelyophilization chamber at approximately the same rate. The freezing ofthe liquid solution from the top and the bottom can be determined bymeasuring the temperature of the solution during the freezing process.The temperature can be measured by inserting at least two thermocouplesinto a vial containing the solution. A first thermocouple 118 can bepositioned at the bottom of the solution, at about the center of thevial, for example, and a second thermocouple 119 can be positioned atthe top of the solution, just below the surface of the solution, inabout the center of the vial, for example. Once a freezing cycle hasbeen optimized for a combination of liquid solution, containerconfiguration, and lyophilization chamber, then in subsequent processingof additional batches temperature monitoring of the containers (e.g.vials) is not necessary.

To freeze the liquid solution from the top and the bottom surfaces atapproximately the same rate, the thermal insulator (e.g., type andthickness) can be selected to provide a water-ice conversion index valuein a range of about −2° C. to about 2° C., or about −1° C. to about 1°C., and/or about −0.5° C. to about 0.5° C. Preferably, the water-iceconversion index is zero or a positive value. The water-ice conversionindex is determined by a method including first plotting thetemperatures reported by the thermocouples at the top (T_(t)) and at thebottom (T_(b)) of the solution as a function of time. The water-iceconversion index is the area between the curves, in ° C.·minute, betweena first nucleation event and the end of water-ice conversion divided bythe water-ice conversion time, in minutes. The water-ice conversion timeis the time necessary for the temperature at the top (T_(t)) of thesolution to reduce in value below the freezing point plateau for thesolution.

The temperature data are collected by loading solution-filled vials intoa lyophilization chamber. The lyophilization tray, at t=0 min, is thencooled to about −60° C. The temperature can then be recorded until atime after which the top and the bottom of the solution cool to atemperature below the freezing point plateau.

The areas, positive and negative, are measured from the first nucleationevent (observable in the plot of temperatures, e.g., such as in FIG. 6)122 until both temperature values cool below the freezing point plateau123. The sum of these areas provides the area between the curves. Whencalculating the area between the curves, the value is positive when thetemperature at the bottom of the vial (T_(b)) is warmer than thetemperature at the top of the vial (T_(t)) 120 and the value is negativewhen the temperature at the top of the vial (T_(t)) is warmer than thetemperature at the bottom of the vial (T_(b)) 121. Preferably, thewater-ice conversion index is zero or a positive value. This conditionwill prevent the consequence that the freezing rate at the bottom of thesolution is significantly higher than that at the top of the solution.Thus, for example, the water-ice conversion index value in one type ofembodiment will be in a range of about 0° C. to about 2° C., or about 0°C. to about 1° C., or about 0° C. to about 0.5° C. For a particularsolution and container configuration, the cooling rate, temperature ofthe tray, and the thermal insulator can be optimized to provide an areabetween the curves at or near 0° C.·minute. For example, FIG. 7 showsthe water-ice conversion indices for 5 wt. % aqueous solutions ofsucrose in vials on a stainless steel tray as a function of the distancefrom the heat sink surface to the stainless steel tray, with the thermalinsulator 108 comprising air within a gap between the heat sink surfaceand the bottom of the stainless steel tray. The tray had a thickness ofabout 1.2 mm.

Still another embodiment of the invention is a lyophilized cake made bya method disclosed herein. The lyophilized cake can include asubstantially dry lyophilized material and a plurality of pores in thelyophilized material having substantially the same pore size. In oneembodiment, the lyophilized cake has a pore size that is substantiallylarger than the pore size of a reference lyophilized cake comprising thesame material as the lyophilized cake but made by a standardlyophilization process (e.g., placing a vial 106 comprising a liquidsolution onto a heat sink 101 within a lyophilization chamber, excludinga thermal insulator between the vial and the heat sink 101, lowering thetemperature of the heat sink 101 and thereby freezing the liquidsolution, and then lyophilizing the frozen solution). Thecross-sectional area of the cylindrical pores of the lyophilized cake ispreferably at least 1.1, 2, and/or 3 times greater than thecross-sectional area of the reference lyophilized cake. In anotherembodiment the lyophilized cake has a substantially consistent pore sizethroughout the cake.

The size of pores in the lyophilized cake can be measured by a BETsurface area analyzer. The effective pore radius (r_(e)), a measure ofthe pore size, can be calculated from the measured surface area of thepores (SSA) by assuming cylindrical pores. The effective pore radiusr_(e) can be determined by the equation r_(e)=2ε/SSA·ρ_(s)·(1−ε) whereSSA is the surface area of the pores, E is the void volume fraction orporosity (ε=V_(void)/V_(total)=n·r_(e) ²/V_(total)), (1−ε) is the soluteconcentration in the volume fraction units, and ρ_(s) is the density ofthe solid.

EXAMPLES

The following examples are provided to illustrate the invention, but arenot intended to limit the scope thereof.

Example 1 Effect of Gap Freezing on Lowering Product Temperature and onPore Enlargement

The effect of gap freezing on the pore enlargement for a lyophilized 10%aqueous sucrose solution was studied. Multiple 20 mL Schott tubing vialswere filled with 7 mL of a 10% aqueous solution of sucrose. These filledvials were placed in a LyoStar II™ (FTS SYSTEMS, INC. Stone Ridge, N.Y.)freeze dryer either directly in contact with a top shelf (heat sinksurface) or on a 6 mm gapped tray. See e.g., FIG. 1. Multiple probedvials were produced by inserting two thermocouples into the solutions,one at the bottom-center of the vial and the other one about 2 mm belowthe liquid surface. See. FIG. 5. The filled vials were then lyophilizedby the following procedure:

1) the shelf was cooled to 5° C. and held at this temperature for 60minutes; next

2) the shelf was cooled to −70° C. and held at this temperature for 200minutes (the internal temperatures of the thermocouple-containing vialswere recorded during freezing);

3) after freezing, the 6 mm gapped tray was removed and these vials wereplaced directly on the bottom shelf (this provided the vials on the topand bottom shelves with the same shelf heat transfer rate duringlyophilization, and thereby a direct comparison of the effect ofdifferent freezing methods could be performed); next

4) the lyophilization chamber was evacuated to a set-point of 70 mTorr,and

5) a primary drying cycle, during which time the internal temperaturesof the frozen samples were recorded, was started. The primary dryingcycle involved (a) holding the samples for 10 minutes at −70° C. and 70mTorr, then (b) raising the temperature at a rate of 1° C./min to −40°C. while maintaining 70 mTorr, then (c) holding the samples for 60minutes at −40° C. and 70 mTorr, then (d) raising the temperature at arate of 0.5° C./min to −25° C. while maintaining 70 mTorr, and then (e)holding the samples for 64 hours at −25° C. and 50 mTorr;

6) a secondary drying followed, and involved raising the temperature ata rate of 0.5° C./min to 30° C. and 100 mTorr, and then holding thesamples for 5 hours at 30° C. and 100 mTorr.

The average product temperatures for the frozen samples in vials on thetop and bottom (gapped-tray) shelves, during primary drying, arepresented in FIG. 8. It can be seen that the temperature profile of thesamples on the bottom shelf is much lower than that of those on the topshelf, which implies that the pore size in the dry layer of the bottomshelf samples is much larger than those on the top shelf, due to theeffect of “gap-freezing.” Theoretically, the temperatures are differentfrom the set point temperatures due to evaporative cooling and/or theinsulative effect of larger pore sizes.

The effective pore radius, r_(e), for the individual lyophilized cakeswas determined by a pore diffusion model. See Kuu et al. “Product MassTransfer Resistance Directly Determined During Freeze-Drying UsingTunable Diode Laser Absorption Spectroscopy (TDLAS) and Pore DiffusionModel.” Pharm. Dev. Technol. (available online at:http://www.ncbi.nlm.nih.gov/pubmed/20387998 and later published in Vol.16, no. 4, p. 343-357, 2011) and incorporated herein. The results arepresented in FIG. 9, where it can be seen that the pore radius of thecakes on the bottom shelf is much larger than that on the top shelf. Theresults demonstrate that the 6 mm gapped tray is very effective for poreenlargement.

Example 2 Acceleration of Drying Rate by Removing Gap Following Freezing

An alternative lyophilization procedure was developed to increase therate of freeze-drying by removing the gap between heat sink shelf andcontainer-loaded shelf following freezing.

Multiple 20 mL Schott tubing vials were filled with 5 mL of a 5% (w/v)aqueous solution of sucrose. Two trays containing these filled vialswere placed in a LyoStar II™ (FTS SYSTEMS, INC. Stone Ridge, N.Y.)freeze dryer in the upper and lower portions of the chamber. The trayswere separated from contact with the heat sink shelves by a thermalinsulator comprising a spacer made of plastic tubing placed on each heatsink shelf, to provide a gap of approximately 6.5 mm between each trayand each heat sink shelf.

For monitoring the product temperature on each shelf, two thermocoupleswere placed in center vials and six thermocouples were placed on theedge locations of the shelves, as shown in FIGS. 10 and 11, wherein thenumbers indicate temperature-probed vials.

The shelf temperature (each) was cooled to −70° C., followed by holdingthe shelf at −70° C. for 90 minutes. At this low shelf temperature,cooling of vials can be accelerated, since cooling with a gap isprimarily driven by radiation. The shelf was then heated to −50° C.,followed by holding the shelf at −50° C. for 60 minutes. After thesolution was frozen the shelf temperature was raised to a highertemperature of −50° C. because after complete freezing of the solutionit is not necessary to maintain it at −70° C. for vacuum pulling. Forone of the trays, the thermal insulator 108 was removed prior to vacuumpulling by removing the spacer.

For primary drying: (a) the lyophilization chamber was then evacuated toa set-point of 100 mTorr, (b) the shelf temperature was held at −50° C.(at 100 mTorr) for 30 minutes; (c) the shelf temperature was then rampedto −15° C. (at 100 mTorr) at a rate of 0.5° C./min; and (d) the shelftemperature was then held at −15° C. (at 100 mTorr) until the end ofprimary drying.

For secondary drying: (e) the shelf temperature was ramped to 30° C. (at100 mTorr), at a rate of 0.5° C./min; and (f) the shelf temperature wasthen held at 30° C. (at 100 mTorr) until the end of secondary drying.

Only the temperature profiles of the center vials (TP01, TP02, TP09 andTP10) and the edge vials along the side walls (TP04, TP07, TP12, andTP15) are used for comparison. The product temperature profiles of thecorner vials (TP03, TP05, TP06, TP08, TP11, TP13, TP14, and TP16) arenot representative for a manufacturing scale freeze dryer due to thestrong thermal radiation from the front and back walls to corner vialsin this freeze dryer. The front wall of the LyoStar™ II freeze dryer isacrylic without insulation. The back wall of the chamber has insulation,but the large amount of heat produced by the fluid pump penetratesthrough the insulation and raises the product temperature to someextent.

Comparison for the Approximate Drying Time for Vials on the to ShelfVersus Vials on the Bottom Shelf

As shown in FIGS. 12 and 13, the drying time on the top shelf (FIG. 12,thermal insulator remaining during vacuum pulling) is much longer thanthat on the bottom shelf (FIG. 13, thermal insulator removed beforevacuum pulling) due to the much higher heat transfer rate on the bottomshelf without a thermal insulator. It can be seen from FIG. 12 thatafter approximate 2670 minutes of cycle time, primary drying was stillnot complete, since the temperature in the center vial, TP02 did notmove out of the plateau level during primary drying. On the other hand,FIG. 13 shows that after approximate 1690 minutes of cycle time, primarydrying was complete, since the temperatures in the center vials, TP09and TP10 moved out of the plateau levels of primary drying. Inlettemperatures of the shelf refrigerant fluids are plotted as Tf on eachof FIGS. 12 and 13.

Comparison for the Product Temperatures of Center Vials, TP01 and TP02on the Top Shelf with TP09 and TP10 on the Bottom Shelf

The comparison is shown in FIG. 14, in which the difference between TP02and TP09 is about 1.7° C.

Comparison for the Product Temperatures of Center Vials TP01 and TP02with the Edge Vials TP04 and TP07 on the to Shelf

The comparison is shown in FIG. 15, in which the difference between TP02and TP07 is about 1.5° C.

Comparison for the Product Temperatures of Center Vials TP09 and TP10with the Edge Vials TP12 and TP15 on the Bottom Shelf

The comparison is shown in FIG. 16, in which the difference between TP09and TP12 is only about 0.2° C., which is much smaller than 1.5° C. onthe top shelf.

Using the same lyophilization cycle, the vials on the tray without athermal insulator between the tray and shelf during drying are driedmuch faster than those with a thermal insulator between the tray andshelf during drying. The tray without a thermal insulator during dryingalso has the advantage of reducing the temperature difference betweenthe center and edge vials, which could be important for sometemperature-sensitive formulations.

A series of non-limiting embodiments is described in the numberedparagraphs below.

1. A method comprising:

loading a container comprising a liquid solution into a lyophilizationchamber comprising a heat sink; the liquid solution comprising a soluteand a solvent and characterized by a top surface and a bottom surface;

providing a thermal insulator between the container and the heat sink;

lowering the temperature of the heat sink and thereby the ambienttemperature in the lyophilization chamber comprising the container andthermal insulator to a temperature sufficient to freeze the liquidsolution from the top and the bottom surfaces at approximately the sametemperature and form a frozen solution and

altering the thermal insulator during or after the lowering step.

2. The method of the preceding paragraph further comprising reducing theambient pressure in the chamber to lyophilize the frozen solution.

3. The method of any one of the preceding paragraphs, wherein thecontainer comprises a vial.

4. The method of any one of the preceding paragraphs, wherein thelyophilization chamber comprises a plurality of heat sinks.

5. The method of any one of the preceding paragraphs, comprising loadingthe container comprising the liquid solution into the lyophilizationchamber between two parallel heat sinks.

6. The method of any one of the preceding paragraphs, wherein the heatsink comprises a heat sink surface, the container comprises a bottom,and the thermal insulator comprises a gap between the heat sink surfaceand the container bottom.

7. The method of any one of the preceding paragraphs, further comprisingloading the container comprising the liquid solution onto a traysurface; wherein the thermal insulator is disposed between the traysurface and the heat sink.

8. The method of any one of the preceding paragraphs wherein thealtering of the thermal insulator comprises removing the thermalinsulator.

9. In a method of freezing a liquid solution for subsequentlyophilization, the liquid comprising top and bottom surfaces anddisposed in a container, and the container disposed in a lyophilizationchamber comprising a heat sink, the improvement comprising separatingthe container from direct contact with the heat sink to thereby freezethe solution from the top and bottom surfaces at approximately the sametemperature and during or after freezing the solution placing thecontainer in thermal contact with the heat sink during a drying process.

10. A lyophilized cake comprising:

a lyophilized material; and

a plurality of pores in the lyophilized material having substantiallythe same pore size; wherein the lyophilized cake is made by the methodof paragraph 2.

11. The lyophilized cake of the preceding paragraph, wherein the poresize is substantially larger than the pore size of a referencelyophilized cake; the reference lyophilized cake comprising the samematerial as the lyophilized cake but made by a method comprising loadinga container comprising a liquid solution into a lyophilization chambercomprising a heat sink; the liquid solution comprising the material anda solvent; excluding a thermal insulator between the container and theheat sink; lowering the temperature of the heat sink and thereby theambient temperature in the lyophilization chamber comprising thecontainer comprising the liquid solution to a temperature sufficient tofreeze the liquid solution; freezing the liquid solution; andlyophilizing the frozen solution.

The foregoing description is given for clearness of understanding only,and no unnecessary limitations should be understood therefrom, asmodifications within the scope of the invention may be apparent to thosehaving ordinary skill in the art.

What is claimed:
 1. A method comprising: loading containers comprising a liquid solution on to a plurality of trays comprising tray surfaces in a lyophilization chamber, the tray surfaces being disposed between parallel upper and lower heat sinks, wherein the trays are optionally thermal conductors and wherein the liquid solution comprises a solute and a solvent; providing a thermal conduction insulator forming a fixed distance from the container on the tray to the respective lower heat sink surface; and lowering the temperature of the heat sinks and thereby the ambient temperature in the lyophilization chamber comprising the containers, tray surfaces and thermal insulators to a temperature sufficient to freeze the liquid solution and form a frozen solution.
 2. The method of claim 1, wherein the thermal conduction insulator comprises one of air, a gas, or a partial or complete vacuum.
 3. The method of claim 2, wherein the thermal conduction insulator is an air gap under partial vacuum.
 4. The method of claim 1, wherein the thermal conduction insulator comprises a solid material.
 5. The method of claim 4, wherein the rigid thermal conduction insulator has a thickness in a range of about 0.5 mm to about 50 mm.
 6. The method of claim 1, wherein the thermal conduction insulator has a thermal conductivity less than about 0.2 W/mK.
 7. The method of claim 1, wherein the tray has a thermal conductivity less than about 0.2 W/mK.
 8. The method of claim 1, wherein the heat sink comprises a refrigerant conduit in thermal communication with the heat sink surface.
 9. The method of claim 1, wherein the container comprises a vial.
 10. The method of claim 1, further comprising reducing the ambient pressure in the chamber to lyophilize the frozen solution.
 11. The method of claim 1, comprising loading arrays of such containers comprising liquid solution into the lyophilization chamber, each array thus comprising center containers, edge containers, and optionally corner containers, wherein the fixed distance at a center container differs from the fixed distance at an edge container.
 12. The method of claim 11, wherein the thermal conduction insulator comprises one of air, a gas, or a partial or complete vacuum.
 13. The method of claim 12, wherein the thermal conduction insulator is an air gap under partial vacuum.
 14. The method of claim 11, wherein the thermal conduction insulator has a thermal conductivity less than about 0.2 W/mK.
 15. The method of claim 11, further comprising altering the thermal insulator during or after the freezing of the liquid solution.
 16. The method of claim 15, wherein the tray is thermally conductive and wherein the altering step comprises placing the tray and heat sink in thermally-conductive contact with each other.
 17. The method of claim 16, wherein the tray is thermally conductive and wherein the altering step comprises eliminating the fixed distance from the tray at a container to the respective lower heat sink surface.
 18. The method of claim 17, wherein the thermal conduction insulator comprises at least one spacer and the altering step includes removing the at least one spacer.
 19. The method of claim 11, further comprising reducing the ambient pressure in the chamber to lyophilize the frozen solution.
 20. The method of claim 1, comprising loading arrays of such containers comprising liquid solution into the lyophilization chamber, each array thus comprising center containers, edge containers, and optionally corner containers, wherein the thermal conductivity of the portion of the thermal conduction insulator forming a fixed distance from a center container on the tray to the respective lower heat sink differs from the thermal conductivity of the portion of the thermal conduction insulator forming a fixed distance from an edge container on the tray to the respective lower heat sink.
 21. The method of claim 15, wherein the thermal conduction insulator has a thermal conductivity less than about 0.2 W/mK.
 22. A lyophilized cake comprising a lyophilized solute made by the method of claim
 1. 